Wildlife crossing structures (WCS) over or under highways can reduce the risk of wildlife-vehicle collisions and reconnect fragmented habitat by facilitating safe wildlife passage. However, we understand little about the degree to which traffic-related light and noise disturbance may impact WCS use and in turn the mitigation value of the WCS. We evaluated the impact of noise and light at WCS on species presence, vigilance behavior, and nocturnality. We used three or four camera traps per WCS to record mammal species at 26 WCS located across interstate and state highways within California for a period of ~one month between 2017-2019. Noise levels (dBA and dBC) and light intensity (scalar illuminance) were recorded at dusk (8pm-12am) for each structure. To capture baseline noise levels and mammal presence within the surrounding area, eight camera traps and two sound meters were simultaneously positioned ~1km from the WCS. This resulted in 29 mammal species being documented across 246 camera stations for a total of 6,420 trap nights. Camera trap data was processed using the online database Wildlife Observer Network (https://wildlifeobserver.net). We used a combination of logistic regression and generalized linear mixed-effects models to examine the influence of traffic noise and light, traffic volume, human presence, surrounding vegetation, elevation and WCS dimensions on species presence. We found that species presence was significantly lower at underpasses than in the surrounding habitat and was greatest at sites with low light pollution and low human presence. Species responses to noise varied, with certain species (e.g. deer) tolerating a higher threshold of noise at WCS than others. Preliminary results suggest increasing noise and light intensity may result in increased levels of vigilance behavior and a shift toward a greater degree of nocturnality. By using a mechanistic framework to examine the non-lethal impacts of light and noise exposure at WCS, we can identify thresholds of noise and light tolerance, and provide recommendations to agencies on mitigation measures such as noise and light - attenuating screens.